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United States Patent |
6,051,219
|
Kubota
|
April 18, 2000
|
Method for reducing malodor with Bacillus subtilis Kubota
Abstract
The present invention relates to an oral composition containing Bacillus
subtilis Kubota FERM BP-5847 for animals.
The oral administration of Bacillus subtilis Kubota FERM BP-5847 can
deodorize feces and urine from livestock, poultry, fish and the like and
also suppresses the occurrence of diarrhea.
Inventors:
|
Kubota; Toyoaki (2-27-19, Todoroki, Setagaya-ku, Tokyo, JP)
|
Appl. No.:
|
913045 |
Filed:
|
September 5, 1997 |
PCT Filed:
|
March 12, 1997
|
PCT NO:
|
PCT/JP97/00778
|
371 Date:
|
September 5, 1997
|
102(e) Date:
|
September 5, 1997
|
PCT PUB.NO.:
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WO97/33487 |
PCT PUB. Date:
|
September 18, 1997 |
Foreign Application Priority Data
Current U.S. Class: |
424/93.462; 435/29; 435/252.5; 435/264; 435/839 |
Intern'l Class: |
A01N 063/00; C12N 001/20; C12Q 001/02 |
Field of Search: |
424/93.462
435/252.5,29,264,839
|
References Cited
U.S. Patent Documents
4996055 | Feb., 1991 | Kurasawa | 424/442.
|
5540924 | Jul., 1996 | Onishi et al. | 424/93.
|
5549890 | Aug., 1996 | Kubo | 424/93.
|
Foreign Patent Documents |
2154590 | Dec., 1977 | JP.
| |
5143916 | Nov., 1980 | JP.
| |
1-144971 | Jun., 1989 | JP.
| |
1-293195 | Nov., 1989 | JP.
| |
4-05336895 | Dec., 1993 | JP.
| |
7-241169 | Sep., 1995 | JP.
| |
7-258049 | Sep., 1995 | JP.
| |
9733487 | Sep., 1997 | WO.
| |
Primary Examiner: Marx; Irene
Assistant Examiner: Afremova; Vera
Attorney, Agent or Firm: Browdy and Neimark
Claims
I claim:
1. A method for reducing the malodor in feces and urine consisting
essentially of orally administering to livestock an amount effective of a
member selected from the group consisting of cells of Bacillus subtilis
Kubota, FERM BP-5847; a culture of Bacillus subtilis Kubota, FERM BP-5847;
freeze-dried cells of Bacillus subtilis Kubota, FERM BP-5847; and
freeze-dried culture of Bacillus subtilis Kubota, FERM BP-5847, which
results in a decrease in the malodor of the feces and urine produced by
said livestock.
2. The method according to claim 1, wherein the member is administered in a
feed composition.
3. The method according to claim 1, wherein the member is administered in
drinking water.
4. The method according to claim 1, wherein the member is administered in a
feed composition containing freeze-dried cells of Bacillus subtilis
Kubota, FERM BP-5847.
5. The method according to claim 1 wherein the member is freeze-dried.
6. The method according to claim 1 wherein the member is diluted and
prepared in a suspension.
7. The method according to claim 1 wherein the member is mixed with a
carrier.
8. The method according to claim 1 wherein the carrier is starch.
Description
FIELD OF THE INVENTION
The present invention relates to an oral composition for animals; more
specifically, the present invention relates to an oral composition
containing Bacillus subtilis Kubota for animals.
PRIOR ART
A vast amount of feces and urine is discharged from pigs, poultry and other
livestock during feeding in pig-pens, poultry farms and various livestock
houses, from which a very strong unpleasant odor evolves, causing a
serious pollution problem.
A bad odor from feces and urine is generated not only from livestock
farmers but also from pets in individual homes and animals and fish and
shellfish in zoos and aquariums. Therefore, there is a need to deodorize
the odor.
PROBLEM TO BE SOLVED BY THE INVENTION
Taking into account such demands from the industry, it is an object of the
present invention to develop a new system capable of deodorizing a wide
variety of excreta from not only livestock but also from pets and other
animals and birds, efficiently in a simple fashion.
SUMMARY OF THE INVENTION
The present inventor has made investigations from various angles so as to
attain the object. Focusing attention on the current status of using
microorganisms such as yeast, vibrio and activated sludge as the bacteria
for treating sewage, the present inventor has further screened useful
bacterial strains as scum treating bacteria and deodorizing bacteria.
Then, the inventor has successfully separated a bacterial strain from
soil, which can reduce scum when used as a sewage treating bacterium and
has a remarkable deodorizing effect.
It is confirmed that the bacterial strain thus separated bacteriologically
belongs to Bacillus subtilis, but the useful properties thereof such as
scum reduction and deodorizing action cannot be observed in conventional
Bacillus subtilis strains. Therefore, the separated strain has been
regarded as a new bacterial strain of Bacillus subtilis and named Bacillus
subtilis Kubota.
The bacterial strain has been deposited under Bacillus subtilis Kubota FERM
BP-5847 in the National Institute of Bioscience and Human-Technology,
Agency of Industrial Science and Technology, Japan. This deposit is the
deposit transferred on Mar. 3, 1997 from the National deposit No. FERM
P-9643 deposited in Oct. 8, 1987.
Bacillus subtilis Kubota (sometimes referred to as "the bacterial strain"
or "BSK bacterium" hereinbelow) directly sprayed on feces and urine can
efficiently deodorize feces and urine. An extremely useful novel finding
has been obtained unexpectedly that when the bacterial strain is
administered orally, feces and urine with no bad odor are excreted.
Accordingly, the inventor has found not only direct deodorizing effect of
the BSK bacterium to deodorize feces and urine if the bacterium is applied
directly to feces and urine but also so-called indirect deodorizing effect
of the BSK bacterium to deodorize feces and urine when the bacterium is
orally administered, and have continued further investigations. Thus, the
present invention has been attained.
In other words, the present invention is essentially based on the technical
concept of an oral composition containing Bacillus subtilis Kubota for
animals.
The present invention will now be described below in detail.
DETAILED DESCRIPTION OF THE INVENTION
In the present invention, Bacillus subtilis Kubota (BSK bacterium; FERM
BP-5847) is used as the deodorizing micro-organism. As has been described
above, the BSK bacterium is regarded bacteriologically to belong to
Bacillus subtilis, but characteristically the bacterium has an excellent
usefulness such as deodorizing action.
The bacteriological properties are described below.
(a) Morphology
1. Shape and size of the cell
Short and small bacillus (1.2-1.4.times.3.6-3.8 .mu.m).
2. Cellular Polymorphism
No specific polymorphism. Being surrounded with cell membrane and having a
nucleus. Also having spore, characteristically, with septum observed.
3. Presence or Absence of Mobility
Mobile. About ten flagella.
4. Presence or Absence of Spore
Spore present.
5. Gram Staining
Gram positive. Stained blue with FAVOR-GSET (Nissui Pharmaceutical Co.
Ltd.).
6. Acid-fast Property
Stained red with Ziehl-Neelsen staining.
Acid-fast bacterium.
(b) Growth State in Individual Culture Media
Inoculating the bacterium suspender in a physiological saline onto a meat
broth agar plate, for cultivation at 37.degree. C. for 24 hours, a good
growth is obserbed.
The periphery of the cology is of R-type, not a round shape. The color is
opaque and pale yellow.
During sheep blood agar cultivation, heart infusion agar cultivation and
TRYPTO-SOYA-AGAR cultivation, the colonies are of R-type, opaque and pale
yellow with good growth. The growth state in each of the culture media is
as follows;
plain agar medium : (+)
sheep blood agar medium : (+)
heart infusion : (+)
TRYPTO-SOYA (Nissui Pharmaceutical Co. Ltd.) : (+)
Sabouraud : (-)
salt egg agar medium : (-)
Drigalski : (-)
Brain-heart infusion liquid medium: (+).
(c) Physiological Properties
(1) Reduction of nitrate : (+)
(2) VP test (+)
(3) Indole generation : (-)
(4) Hydrogen sulfide generation : (-)
(5) Starch hydrolysis (+)
(6) Citric acid utilization : (+)
(7) Dye generation : water soluble
(8) Urease : (-)
(9) Oxidase (+)
(10) Catalase (+)
______________________________________
(11) Growth acid poor
neutral good
Temperature 37.degree. C.
good
30.degree. C.
good
25.degree. C.
good
(15) Performance in oxygen
aerobic
(16) O-F test F
______________________________________
(17) Generation of acid and gas from sugars
______________________________________
acid gas
______________________________________
L-arabinose (-) (-)
D-xylose (-) (-)
D-glucose (+) (-)
D-fructose (+) (-)
maltose (+) (-)
sucrose (-) (-)
lactic acid (-) (-)
trehalose (.+-.) (-)
D-sorbitol (-) (-)
D-mannitol (-) (-)
D-inositol (-) (-)
starch (-) (-)
(d) Other properties
Esculin decomposition (+)
Cellulose decomposition (+)
Malonic acid utilization
(+)
Arginine decomposition (-)
Lysine decarboxylation (+)
Ornithine decarboxylation
(-)
Phenylalanine deamination
(-)
Hemolytic property .beta.-type hemolysis
Auxotrophy (+)
.beta.-galactosidase (-)
______________________________________
The novel strain Bacillus subtilis Kubota isolated in the present invention
exhibits good growth in any common nutrient medium. Extract solutions from
poultry feces, extract solutions from livestock feces, etc. particularly
serve as an excellent culture broth.
In cultivation at 20 to 40.degree. C., preferably 30 to 38.degree. C. for
10 to 24 hours with or without using an incubator, the strain shows an
excellent growth.
The resulting culture may be used as it is, because the culture contains a
great amount of the cells thereof.
Additionally, the cells may be separated by centrifugation or the like from
the culture, which are then thoroughly washed and used as they are as the
cultivated cells; such cells may be used as a paste form such as wet cake
in the above case or the cells may be further freeze-dried; alternatively,
the cells may be diluted and prepared in suspension, for use in the form
of such a processed product. It is needless to say that the processed
product of the culture may also be used similarly. BSK bacterium
(including the culture and the processed products thereof) of themselves
may be used in the form of oral composition; additionally, the cells may
be formulated into pharmaceutical agents for animals, after addition and
mixing of starch, talc and other routine components into the cells. As the
formulation, powder, granule, table, liquid, paste and other forms may
appropriately be selected.
The oral composition in accordance with the present invention may be
formulated as feed type, and in such a case, the cells of BSK bacterium
may, of course, be added as they are into feeds; besides, the cells may be
prepared preliminarily as a premix, which may be used; or the cells
formulated as the aforementioned pharmaceutical type may De added into
feeds.
Namely, the oral compostion of the present invention may be used as a
pharmaceutical type or a feed type; and the cells of BSK bacterium (after
premixed or formulated if necessary) may directly be administered orally
as they are or they may be added and mixed as a feed additive into feeds
or drinking water prior to intake. For drinking water, the cells of BSK
bacterium may be directly added and mixed into drinking water and
additionally, the cells of BSK bacterium may be packaged in a
water-permeable bag which may be hung in a tank of drinking water.
Furthermore, the cells of BSK bacterium may be possibly mixed with a feed
or a pet food product or a drink product, for preparation as a final feed
product. The amount of the microorganism in a feed varies depending on the
species, age, etc. of a subject animal, but it may satisfactorily be said
as a tentative standard that the BSK bacteria added in feeding water
should be fed ad libtum at 10.sup.7 cells per pig weighing 70 kg per day.
At oral administration tests in mice, the LD.sub.50 of BSK bacterium to be
used in the oral composition of the present invention is nearly infinite;
at coating tests on murine skin, no change has been observed. Hence, no
problem in safety is noticed. Therefore, BSK bacterium can be used safely,
without any adverse effect on animals at other microbial doses above the
dose described previously.
Still furthermore, BSK bacterium has already been tested for safety at the
TOKYO FOOD SANITATION ASSOCIATION, FOOD RESEARCH LABORATORY, which is an
inspection organization authorized by the Minister of Health and Welfare.
In that respect, the safety of the bacterium has been validated.
As has been described above, the composition of the present invention has
prominent characteristics in that the composition orally administered can
deodorize feces indirectly in a sense, in addition to direct application
thereof over feces and urine.
Therefore, the use of the composition simply added into assorted seeds,
other feeds or drinking water and the like, or simple feeding of the
composition as a feed, pet food, liquid, etc. can deodorize feces and
urine. Thus, laborious works can be skipped with the resultant energy
saving; labor can be saved an environment with bad odor in particular,
which is advantageous for not only workers for cattle farmers but also
near-by residents of such cattle farmers because they can be free from the
bad odor. Thus, the present invention is very excellent in terms of
pollution control.
According to the present invention, furthermore, feces and urine from pets
in homes and pet shops, animals at zoos and aquariums, and animals at pet
hotels and veterinary clinics can be deodorized in a simple manner, so bad
odor therefrom can be eliminated from these environment; even if feces and
urine are left to stand alone, the generation of bad odor can be
suppressed. In light of this point, the present invention is prominent.
Still further, the composition of the present invention, if put as it is or
together with feeds in a water tank or a fish preserve, can prevent the
generation of bad odor in the water therein. Therefore, the present
invention is very useful in cleaning up water at aquariums, fish preserves
and fishing ponds, and water in water tanks for tropical fish and other
pets at homes and pet shops, and in preventing the generation of bad odor
or fishy odor.
Furthermore, the composition of the present invention has a good action of
curing intestinal disorder, and when administered orally, the composition
distinctively suppresses the occurrence of diarrhea. Particularly, pigs
are animals readily having loose bowels, so if the occurrence of diarrhea
is suppressed in young pigs, they can make good growth, leading to the
increase of profit. Additionally, chicken and other animals can
dramatically elevate the projection of meat and eggs. From these respects,
the present invention is very excellent.
Examples of the present invention will now be described hereinbelow.
EXAMPLE 1
______________________________________
Starch 10 g
Meat extract 3 g
Peptone 10 g
Sodium chloride 5 g
Water 1,000 ml (pH 7.0)
______________________________________
The medium of the composition described above was placed in a 500-ml flask,
into which was inoculated Bacillus subtilis Kubota FERM BP-5847, for
stationary culture at 37 .degree. C. for 20 hours.
The resulting culture broth was centrifuged to recover the cells,which were
then mixed with 300 g of starch. The resulting mixture was defined as a
microbial formulation.
EXAMPLE 2
Commercially available growing feeds were fed to 50 young pigs at the stage
of growth maintained at a pig farmer. The cells of BSK strain recovered in
Example 1 were added to drinking water to 10.sup.5 cells/day, and the
resulting water was fed ad libitum.
One month later, the odor and ammonia concentration in the pig-pens were
determined. The concentration of the odor was determined by the
three-point comparison method (Notification No. 238 of the Metropolitan
Environmental Pollution Control Ordinate),while the ammonia concentration
was measured by an ammonia detector.
Consequently, the concentration of the odor, which was 117 prior to
feeding, was reduced to 74 after feeding (one month later), which means
odor reduction and elimination by as much as by37%. Alternatively, the
ammonia concentration was 2.3 prior to feeding but 1.0 after feeding (one
month later), which means the ammonia was reduced and eliminated by as
much as 57%. During the growing term, eight of the young pigs had
diarrhea, but one month later, then, no young pig with diarrhea was
observed.
EXAMPLE 3
Twenty dairy cows were divided in two groups, of ten cows, each and then,
green oats as a crude feed was blended with a commercially available feed.
Subsequently, the resulting feed was further blended with the microbial
formulation as prepared in Example 1; the added amount of the microbial
formulation was 0.1%. The resulting mixture was fed to a test lot of 10
dairy cows for one week. On contrast, the feed with no addition of the
microbial formulation was fed to a control lot of 10 dairy cows.
One week later, no specific bad odor was observed in the dairy cow houses
of the test lot, and even in the dairy cow houses wherein feces and urine
were left to stand as they were, milking was performed with no
disturbance. In contrast, unendurable smell filled in the diary cow houses
of the control lot throughout a term of one week. Even in the test lot,
however, the cessation of the microbial feeding caused generation of a bad
smell on the very next day.
EXAMPLE 4
A feed was prepared by adding the freeze-dried product containing starch of
the cells as prepared in Example 1 at 0.2% into a commercially available
goldfish feed, and the feed was used to feed goldfish in a commercially
available round goldfish basin.
Despite no change of water or no cleaning of the goldfish basin for a
month, the water in the goldfish basin was clean, with no occurrence of a
bad odor such as a fishy odor.
Effect of the Invention
A bad odor from a variety of animals including fish can be eliminated by
the oral administration of BSK bacterium and therefore, the composition of
the present invention can be applied widely not only to fish, cattle and
poultry in homes and of cattle farmers and fish breeders but also to pets,
experimental animals and animals at zoos and aquariums. Hence, the
composition serves very well for pollution control of a bad odor currently
drawing serious concern, which enables the establishment of a livestock
industry in the suburbs of cities and also serves markedly for livestock
farmers in the suburbs of cities. Thus, the composition contributes much
for the propagation of the domestic livestock industry.
Reference to the Microorganism Deposited under the Provision of Article
13-2
1. Bacillus subtilis Kubota
a. Name and address of the depository at which the microorganism was
deposited;
name: The National Institute of Bioscience and Human-Technology, Agency of
Industrial Science and Technology, Ministry of International Trade and
Industry
address: 1-3, Higashi 1 chome, Tsukuba-shi, Ibaraki-ken, 305, Japan
b. Deposition date at the depository (a) Oct. 8, 1987
c. Accession Number of the deposition, designated by the depository (a)
FERM BP-5847
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